PDF(Pubmed)
Abstract：
OBJECTIVE: In gout, monosodium urate crystals are taken up by macrophages, triggering the activation of the NLRP3 inflammasome and the maturation of IL-1β. This study aimed to investigate the role of integrin CD11b in inflammasome activation in macrophages stimulated by MSU.
METHODS: BMDM from WT and CD11b KO mice were stimulated in vitro with MSU crystals. Cellular supernatants were collected to assess the expression of the inflammatory cytokines by enzyme-linked immunosorbent assay and western blot methods. The role of integrin CD11b in MSU-induced gouty arthritis in vivo was investigated by intra-articular injection of MSU crystals. Real-time extracellular acidification rate and oxygen consumption rate of BMDMs were measured by Seahorse Extracellular Flux Analyzer.
RESULTS: We demonstrate that CD11b-deficient mice developed exacerbated gouty arthritis with increased recruitment of leukocytes in the joint and higher IL-1β levels in the sera. In macrophages, genetic deletion of CD11b induced a shift of macrophage metabolism from oxidative phosphorylation to glycolysis, thus decreasing the overall generation of intracellular ATP. Upon MSU stimulation, CD11b-deficient macrophages showed an exacerbated secretion of IL-1β. Treating wild-type macrophages with a CD11b agonist, LA1, inhibited MSU-induced release of IL-1β in vitro and attenuated the severity of experimental gouty arthritis. Importantly, LA1, was also effective in human cells as it inhibited MSU-induced release of IL-1β by peripheral blood mononuclear cells from healthy donors.
CONCLUSIONS: Our data identified the CD11b integrin as a principal cell membrane receptor that modulates NLRP3 inflammasome activation by MSU crystal in macrophages, which could be a potential therapeutic target to treat gouty arthritis in human patients.
METHODS: BMDM from WT and CD11b KO mice were stimulated in vitro with MSU crystals. Cellular supernatants were collected to assess the expression of the inflammatory cytokines by enzyme-linked immunosorbent assay and western blot methods. The role of integrin CD11b in MSU-induced gouty arthritis in vivo was investigated by intra-articular injection of MSU crystals. Real-time extracellular acidification rate and oxygen consumption rate of BMDMs were measured by Seahorse Extracellular Flux Analyzer.
RESULTS: We demonstrate that CD11b-deficient mice developed exacerbated gouty arthritis with increased recruitment of leukocytes in the joint and higher IL-1β levels in the sera. In macrophages, genetic deletion of CD11b induced a shift of macrophage metabolism from oxidative phosphorylation to glycolysis, thus decreasing the overall generation of intracellular ATP. Upon MSU stimulation, CD11b-deficient macrophages showed an exacerbated secretion of IL-1β. Treating wild-type macrophages with a CD11b agonist, LA1, inhibited MSU-induced release of IL-1β in vitro and attenuated the severity of experimental gouty arthritis. Importantly, LA1, was also effective in human cells as it inhibited MSU-induced release of IL-1β by peripheral blood mononuclear cells from healthy donors.
CONCLUSIONS: Our data identified the CD11b integrin as a principal cell membrane receptor that modulates NLRP3 inflammasome activation by MSU crystal in macrophages, which could be a potential therapeutic target to treat gouty arthritis in human patients.
摘要：
目标：在痛风中，尿酸单钠晶体被巨噬细胞吸收，触发NLRP3炎性体的激活和IL-1β的成熟。本研究旨在探讨整合素CD11b在MSU刺激的巨噬细胞炎症小体激活中的作用。
方法:用MSU晶体体外刺激来自WT和CD11bKO小鼠的BMDM。收集细胞上清液以通过酶联免疫吸附测定和蛋白质印迹方法评估炎性细胞因子的表达。通过关节内注射MSU晶体研究了整合素CD11b在体内MSU诱导的痛风性关节炎中的作用。通过海马胞外通量分析仪测量BMDMs的实时细胞外酸化速率和耗氧率。
结果：我们证明CD11b缺陷小鼠发展为痛风性关节炎，关节中白细胞募集增加，血清中IL-1β水平升高。在巨噬细胞中，CD11b的遗传缺失诱导巨噬细胞代谢从氧化磷酸化转变为糖酵解，从而减少细胞内ATP的整体生成。在MSU刺激时，CD11b缺陷型巨噬细胞显示IL-1β分泌加剧。用CD11b激动剂治疗野生型巨噬细胞，LA1在体外抑制MSU诱导的IL-1β释放，并减轻实验性痛风性关节炎的严重程度。重要的是,LA1在人细胞中也有效，因为它抑制了MSU诱导的健康供体外周血单核细胞释放的IL-1β。
结论：我们的数据确定CD11b整合素是一种主要的细胞膜受体，通过巨噬细胞中的MSU晶体调节NLRP3炎性体的活化，这可能是治疗人类痛风性关节炎的潜在治疗靶点。
方法:用MSU晶体体外刺激来自WT和CD11bKO小鼠的BMDM。收集细胞上清液以通过酶联免疫吸附测定和蛋白质印迹方法评估炎性细胞因子的表达。通过关节内注射MSU晶体研究了整合素CD11b在体内MSU诱导的痛风性关节炎中的作用。通过海马胞外通量分析仪测量BMDMs的实时细胞外酸化速率和耗氧率。
结果：我们证明CD11b缺陷小鼠发展为痛风性关节炎，关节中白细胞募集增加，血清中IL-1β水平升高。在巨噬细胞中，CD11b的遗传缺失诱导巨噬细胞代谢从氧化磷酸化转变为糖酵解，从而减少细胞内ATP的整体生成。在MSU刺激时，CD11b缺陷型巨噬细胞显示IL-1β分泌加剧。用CD11b激动剂治疗野生型巨噬细胞，LA1在体外抑制MSU诱导的IL-1β释放，并减轻实验性痛风性关节炎的严重程度。重要的是,LA1在人细胞中也有效，因为它抑制了MSU诱导的健康供体外周血单核细胞释放的IL-1β。
结论：我们的数据确定CD11b整合素是一种主要的细胞膜受体，通过巨噬细胞中的MSU晶体调节NLRP3炎性体的活化，这可能是治疗人类痛风性关节炎的潜在治疗靶点。
